The goal of the proposed research is to determine the mechanisms underlying failure of action potentials to invade every axon branch in the presynaptic terminal arborization. Intracellular Ca++ may regulate axon excitability. Using ion-selective electrodes and intracellular recording techniques, the extent of intracellular Ca++ accumulation, the effects of injecting Ca++ or EGTA, the relative contribution of Ca++ currents during repetitive action potentials, and the amount of intracellular Na+ accumulation under conditions known to cause conduction block will be measured. Axonal adenylate concentrations increase during repetitive stimulation and may also affect conduction. Using sensitive biochemical techniques, the amounts of adenylate and adenosine released by nerve and muscle and the amounts taken up by the nerve will be assayed. The effects of these compounds on axon excitability will be tested using patch-clamp recording techniques. Recent experiments have shown clearly that action potentials do not propagate to the terminal region of motor axons at the mouse neuromuscular junction; the terminal portions are depolarized due to passive current invasion beyond a more proximal site of conduction block. Using Nomarski optics to visualize terminal branches, terminal depolarization and synaptic responses will be recorded with patch-clamp electrodes to determine if the site of blockage varies and whether transmitter release by individual terminal branches is graded or all-or-none. To interpret the significance of any alterations in terminal excitability or passive depolarization, the extent of the synaptic contact zone in the terminals will be determined from serial sections obtained with the electron microscope. These studies should elucidate the underlying mechanisms and the extent of variable nerve terminal invasion by the action potential; this is a fundamental issue, for variable terminal activation could modulate the divergence of information throughout the nervous system and the effectiveness of single synapses.